Antiviral guanine derivatives

ABSTRACT

A compound of formula (I)                    
     or a salt or acyl derivative thereof, in which X represents chlorine, C 1-6  alkoxy, phenoxy, phenyl C 1-6  alkoxy, NH 2 , —OH or —SH, is useful in treating viral infections.

This is a continuation of application Ser. No. 07/847,833, filed Mar. 9,1994, now abandoned, which is a continuation of 07/697,853, filed May 9,1991, abandoned, which is a continuation application of application Ser.No. 07/085,216, now U.S. Pat. No. 5,075,445 filed on Aug. 12, 1987,which is a Continuation application of application Ser. No. 06/641,300filed on Aug. 16, 1984 abandoned.

The present invention relates to compounds having antiviral activity,processes for their preparation and pharmaceutical compositionscontaining them.

The compound 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine of formula (A)

is disclosed in Synthetic Communications, 2(6), 345-351 (1972) but nopharmaceutical activity has been indicated for the compound in this orany other published document. We have repeated the synthesis of thecompound as described in the above publication, and have shown that theproduct is a mixture of the compound of formula (A), its monobenzylether and its dibenzyl ether, this mixture having a melting point and uvspectrum in agreement with those reported in the publication for thesupposedly ‘pure’ compound of formula (A). Our analysis of the productproduced by the above synthesis showed that it contained 45-50% byweight of the compound of formula (A), 45-50% by weight of themonobenzyl ether and 5% or less by weight of the dibenzyl ether.

By different synthetic routes, we have prepared the compound of formula(A) in a substantially pure form and have found that it has anti-viralactivity. This activity is also shown by certain derivatives of thecompound of formula (A).

According to the present invention there is provided a compound offormula (I)

or a salt, phosphate ester or acyl derivative thereof, in which Xrepresents chlorine, straight or branched chain C₁₋₆ alkoxy, preferablymethoxy, phenoxy, phenyl C₁₋₆ alkoxy, —NH₂, —OH or —SH with the provisothat, when X is —OH, the compound of formula (I) is in a purity state ofgreater than 50% by weight of pure compound.

The term ‘acyl derivative’ is used herein to include any derivative ofthe compounds of formula (I) in which one or more acyl groups arepresent. Such derivatives include biological precursors of the compoundsof formula (I) in addition to those derivatives which are per sebiologically active.

Examples of acyl derivatives of the compounds of formula (I) are thosewherein one or both of the hydrogens in the acyclic OH groups, and/orone of the hydrogen atoms in the —NH₂ group, are replaced by

wherein R is hydrogen or an alkyl, aryl, aralkyl or heterocyclyl group.

Examples of alkyl groups R include straight and branched chain groupscontaining up to 18 carbon atoms, preferably up to 6 carbon atoms.Particular examples are methyl, ethyl, t-butyl and pentyl.

Examples of aryl groups R include phenyl optionally substituted with upto five preferably up to three groups.

Examples of aralkyl groups R include phenyl-C₁₋₆ alkyl groups such asbenzyl.

Examples of heterocyclyl groups R include single or fused ringscontaining one or two hetero-atoms in each ring, selected from oxygen,nitrogen and sulphur.

Examples of phosphate esters of the compounds of formula (I) includethose where one or both of the acyclic —OH groups are replaced by

or salts thereof, or where the two —OH groups are replaced by a bridging

group

Salts, phosphate esters and acyl derivatives of the compounds of formula(I) are preferably pharmaceutically acceptable, but non-pharmaceuticallyacceptable compounds are also within the scope of the present invention,since these are useful as intermediates in the preparation ofpharmaceutically acceptable compounds.

The compounds of formula (I) are defined herein as including tautomersof formula (I), wherein the —OH and —SH substituents are replaced by ═Oand ═S substituents respectively.

A particular group of compounds of the invention are those of formula(II)

or pharmaceutically acceptable salts thereof, in which X is as definedin formula (I), and each of R¹, R² and R³ represents hydrogen or an acylgroup of formula

in which R⁴ is C₁₋₁₈ alkyl or imidazolyl, or R¹ or R² represents aphosphate ester group of formula

or R¹ and R² together represent a

bridging group.

Subject to the aforementioned purity proviso in relation to compounds ofthe invention, a preferred compound of the present invention is thecompound of formula (A)

or a salt or acyl derivative therof.

In a further aspect of the invention there is provided a compound offormula (A) in a purity state of greater than 60% preferably greaterthan 80% more preferably greater than 90% and particularly preferablymore than 95% by weight of pure compound.

In yet a further aspect of the invention, there is provided an isolated,substantially completely pure compound of formula (A), or apharmaceutically acceptable salt thereof.

The invention also provides a compound of formula (A) in crystallineform having a melting point of 275-277° C.

The compounds of the present invention have antiviral activity, and arepotentially useful in the treatment of infections caused by herpesviruses, such as herpes simplex type 1, herpes simplex type 2 andvaricella zoster viruses.

Accordingly, the present invention also provides a compound of formula(I) or a pharmaceutically acceptable salt, phosphate ester or acylderivative thereof, for use as an active therapeutic substance, and inparticular for use in the treatment of viral infections. In this aspectof the invention, the compounds of formula (I) are not subject to theaforementioned purity proviso.

Examples of pharmaceutically acceptable salts of the compounds offormula (I) are those formed with organic bases, preferably with aminessuch as ethanolamines or diamines; and alkali metals, such as sodium andpotassium; and acid addition salts formed with a pharmaceuticallyacceptable acid such as hydrochloric acid, orthophosphoric acid andsulphuric acid.

The compound of formula (A) or a salt thereof may be prepared byconverting the group X in a compound of formula (III).

in which X, excluding —OH, is as defined in formula (I); R^(a) andR^(b), which may be the same or different, are each hydrogen or O—protecting groups, preferably acyl groups; and Y is chlorine or—NHR^(c), in which R^(c) is hydrogen or acyl, to an —OH group by meansof hydrolysis, preferably acid hydrolysis, when X is other than NH₂, or,when X is —NH₂, by means of a deaminase reaction, or when Y is chlorineand X is —OH, converting Y to a —NH₂ group by reaction with ammoniaunder pressure in accordance with known methods, and subsequently, ifdesired, converting the compound of formula (A) to a salt thereof bytreatment with an acid or base.

Acyl groups R^(a), R^(b) and R^(c) may be those of formula

as hereinbefore defined.

Examples of groups R^(a) and R^(b) in formula (III) are acetyl andcyclic acetal such as isopropylidene. R^(c) is preferably acetyl orhydrogen.

A preferred process for preparing the compound of formula (A) comprisestreating a compound of formula (III) in which X is chlorine, Y is —NH₂and R^(a) and R^(b) are each acetyl, with aqueous mineral acid,preferably hydrochloric acid.

Compounds of formulae (III) when each of R^(a), R^(b) and R^(c) ishydrogen or acyl, are themselves compounds of the invention, having theadditional utility as intermediates for the preparation of the compoundof formula (A).

In a further aspect of the invention, compounds of formula (I) or acylderivatives thereof, together with, compounds of formula (III), may beprepared by treating a compound of formula (IV).

in which X is as defined in formula (I) and Y is as defined in formula(III), with a compound of formula (V)

in which R^(a) and R^(b) are as defined in formula (III) and Z is aleaving group such as Cl, Br, or I, preferably Br.

Compounds of formula (IV) are either known compounds or can be made fromknown compounds by known methods.

Compounds of formula (V) in which Z is bromine may be prepared bybrominating a compound of formula (VI).

preferably by treatment with carbon tetrabromide and triphenylphosphinein an organic, aprotic solvent, such as dimethylformamide.

Compounds of formula (V) in which Z is Cl or I may be prepared in ananalogous manner.

Compounds of formula (VI) in which R^(a) and R^(b) are identical may beprepared according to the following schematic process.

Acyl derivatives of compounds of formula (I) may also be prepared byacylating an optionally protected compound of formula (I) in accordancewith conventional acylating processes known in the art, and wherenecessary, deprotecting the resulting product.

The acylation reaction may be carried out by using an acylating agentcontaining a

wherein R is as hereinbefore defined.

In a particular aspect of this process, the acylating agent contains the

in which R⁴ is C₁₋₁₈ alkyl, or is N,N′-carbonyldiimidazole.

Examples of acylating agents suitable for the above process arecarboxylic acids, acid halides or acid anhydrides, preferably anhydridesor acids.

When the acylating agent is a carboxylic acid, a coupling agent such asdicyclohexylcarbodiimide should be included, but this is not necessarywhen the acylating agent is an acid anhydride.

The acylation reaction may produce a single acyl derivative of acompound of formula (I), or a mixture of derivatives, depending on anumber of factors, such as the relative amounts and chemical natures ofreactants, the physical conditions of the reaction, and the solventsystem. Any mixture produced in this way may be separated into its purecomponents using standard chromatographic techniques.

The above described acylation process of the invention can yield mono-,di-, or tri-acylated derivatives of compounds of formula (I) accordingto the form of protection/deprotection utilised. The following areexamples of products obtained by different methods:

(a) Di-acylated derivatives of the two acyclic-OH groups may be obtainedby direct acylation of unprotected compounds of formula (I) or acylationof protected intermediates of compounds of formula (I) in which the —NH₂group is protected by, for example, a monomethoxytrityl group, andsubsequent deprotection by treatment with acid.

(b) Mono-acylated derivatives of one of the acyclic —OH groups may beobtained by acylation of protected intermediates of compounds of formula(I) in which the —NH₂ group and the other acyclic —OH group are bothprotected by, for example, monomethoxytrityl groups, and subsequentdeprotection by acid treatment.

(c) Mono-acylated derivatives of the NH₂ group may be obtained byacylation of protected intermediates of compounds of formula (I) inwhich both acyclic —OH groups are protected by, for exampletrimethylsilyl groups, and subsequent deprotection.

The various protected intermediates of compounds of formula (I) may beprepared in accordance with standard procedures by, for example,treatment of the compounds with monomethoxytrityl chloride (forprocesses (a) and (b)) or with chlorotrimethylsilane (for process (c)).

Protected intermediates of compounds of formula (I) may also be used toprepare phosphate esters of the compounds.

Accordingly, in a further process aspect of the invention, there isprovided a process for preparing a mono-phosphate ester of a compound offormula (I) which comprises treating a protected intermediate of thecompound of formula (I) in which one of the acyclic —OH groups and the—NH₂ group are protected, preferably by monomethoxytrityl groups, withcyano ethyl phosphoric acid and subsequently deprotecting the resultantproduct by treatment with acid, preferably acetic acid.

If desired, the reaction product after treatment with cyano ethylphosphoric acid is treated with aqueous ammonia, which yields theammonium salt of the phosphate ester as the final product.

Compounds of formula (I) or salts thereof may also be prepared byhydrolysing the 1,3-dioxane ring of a compound of formula (VII).

in which X is as defined in formula (I) and R^(c) is as defined informula (III), provided that R^(c) is not acyl when X is other than OH,and subsequently, if desired, converting the compound of formula (I)thus formed to a salt by treatment with an acid or base.

When R^(c) is an acyl group, a basic N-deprotection step is required toform the compound of formula (A). This can be carried out prior to orafter hydrolysis by treatment with, for example, (i) a solution of NaOMein CH₃OH or (ii) a solution of NH₃ in CH₃OH.

Preferably the hydrolysis of compounds of formula (VII) is carried outin acid medium. The compounds of formula (VII) in which X is alkoxy,phenoxy, phenylalkoxy or —SH are conveniently prepared in situ byreacting the compound of formula (VII) in which X is chlorine with anadditional reactant containing an X¹ substituent, wherein X¹ is alkoxy,phenoxy, phenylalkoxy or sulphur. These intermediates can then behydrolysed to compounds of formula (I) without isolating them from thereaction mixture.

The additional reactant containing the X¹ moiety may be a sodiumalkoxide, phenoxide or phenylalkoxide, or sodium hydrosalphide (when X¹is sulphur).

Acid hydrolysis of a compound of formula (VII) in which X is chlorinewill yield a compound of formula (I) in which X is chlorine, or acompound of formula (A) depending on acid strength and reactionconditions.

For example, treatment of the compound of formula (VII) in which X ischlorine with dilute HCl (1.0M) at 60° C. for 24 hours or with 2 M HClunder reflux for 1.5 hours, will yield the compound of formula (A).Treatment of the same compound of formula (VII) with 2 M HCl intetrahydrofuran at room temperature will yield the compound of formula(I) in which X is chlorine.

If desired, the compound of formula (VII) in which X is chlorine may beconverted to the compound of formula (VII) in which X is amino, prior toacid hydrolysis.

The conversion may be achieved by treatment with sodium azide indimethylformamide to form an azido intermediate in which X is replacedby an azide moiety, followed by reduction of the intermediate withammonium formate/palladium-on-charcoal in methanol.

The intermediate compound of formula (VII) in which X is chlorine andR^(c) is hydrogen may be prepared by treating a compound of formula(VIII).

with a compound of formula (IX)

The reaction may be carried out in an inert organic solvent, preferablydimethylformamide, in the presence of an inorganic base, preferablypotassium carbonate.

The compound of formula (VIII) may itself be prepared by brominating acompound of formula (X)

The reaction is preferably carried out by treating the compound offormula (X) with carbon tetrabromide and triphenylphosphine in anorganic, aprotic solvent such as dimethylformamide.

The compound of formula (X) may itself be prepared by treating acompound of formula (XI)

with 2,2-dimethoxypropane and p-toluenesulphonic acid in the presence ofacetone or tetrahydrofuran.

The compounds of formulae (IX) and (XI) are known compounds or can beprepared from known compounds by known methods.

The compounds of formulae (VII), (VIII) and (X) are novel intermediatesand as such form further aspects of the present invention.

A compound of formula (I) or pharmaceutically acceptable salt, phosphateester or acyl derivative thereof may be formulated for use in apharmaceutical composition. Accordingly, in a further aspect of theinvention, there is provided a pharmaceutical composition whichcomprises a compound of formula (I) or pharmaceutically acceptable salt,phosphate ester or acyl derivative thereof together with apharmaceutically acceptable carrier or excipient.

A composition which may be administered by the oral route to humans maybe compounded in the form of a syrup, tablet or capsule. When thecomposition is in the form of a tablet, any pharmaceutical carriersuitable for formulating such solid compositions may be used, forexample magnesium stearate, starch, lactose, glucose, rice, flour andchalk. The composition may also be in the form of an ingestible capsule,for example of gelatin, to contain the compound, or in the form of asyrup, a solution or a suspension. Suitable liquid pharmaceuticalcarriers include ethyl alcohol, glycerine, saline and water to whichflavouring or colouring agents may be added to form syrups. Thecompounds may also be presented with a sterile liquid carrier forinjection.

The composition may also be formulated for topical application to theskin or eyes.

For topical application to the skin, the composition may be in the formof a cream, lotion or ointment. These formulations may be conventionalformulations well known in the art, for example, as described instandard books of pharmaceutics and cosmetics, such as Harry'sCosmeticology published by Leonard Hill Books and the BritishPharmacopaeia.

The composition for application to the eyes may be a conventionaleye-drop composition well known in the art, or an ointment composition.

Preferably, the composition of this invention is in unit dosage form orin some other form that the patient may administer to himself a singledose. A suitable dosage unit might contain from 50 mg to 1 g of activeingredient, for example 100 to 500 mg. Such doses may be administered 1to 4 times a day or more usually 2 or 3 times a day. The effective doseof compound will in general be in the range of from 1.0 to 20 mg/kg ofbody weight per day or more usually 2.0 to 10 mg/kg per day.

In a further aspect of the invention there is provided a method oftreating viral infections in a human or non-human animal, whichcomprises administering to the animal an effective, non-toxic amount ofa compound of formula (I) or a pharmaceutically acceptable salt,phosphate ester or acyl derivative thereof.

The preparation of compounds of the invention is illustrated by thefollowing Examples.

EXAMPLE 1 5-(2-Hydroxyethyl)-2,2-dimethyl-1,3-dioxan

To a suspension of lithium aluminium hydride (2.87 g, 76 mmol) intetrahydrofuran (125 ml), a solution of triethyl1,1,2-ethanetricarboxylate (9.2 ml, 9.85 g, 40 mmol) in tetrahydrofuran(25 ml) was added dropwise with stirring over 2 hours. Excess reagentwas then quenched with aqueous tetrahydrofuran (1:2). The inorganicsalts were filtered off and washed with ethanol (100 ml). The filtrateand washings were combined and the solvent was evaporated under reducedpressure to afford a colourless oil (4.85 g). To a suspension of thisoil in acetone (100 ml.), 2,2-dimethoxypropane (25 ml) andp-toluenesulphonic acid monohydrate (2.3 g, 12 mmol) were added and themixture was stirred for 1 hour. The resulting solution was neutralisedwith Amberlite IR 45 (OH⁻ form, methanol washed), filtered and thesolvent evaporated under reduced pressure. The residue was purified bycolumn chromatography on silica gel, eluting with chloroform-methanolmixtures (40:1 and 25:1) to afford5-(2-hydroxyethyl)-2,2-dimethyl-1,3-dioxan as a colourless liquid (3.01g, 47%); νmax (film) 3420, 2940, 1375, 1200 and 1080 cm⁻¹; δ_(H) (CDCl₃)1.34-1.70 (8H, m, C(CH₃)₂ and CH₂CH₂OH), 1.7-2.1 (1H, m, CH), 2.15 (1H,br, D₂O exchangeable, OH), and 3.5-4.0 (6H, m, 3×CH₂O); (Found: C,58.33; H, 10.11%. C₈H₁₆O₃ 0.25H₂O requires C, 58.34; H, 10.10%. [M−CH₃]⁺found 145.0864; C₇H₁₃O₃ requires 145.0865).

Alternative Procedure for Preparation of5-(2-Hydroxyethyl)-2,2-dimethyl-1,3-dioxan (Example 1)

A. 1,4-Dihydroxy-2-hydroxymethylbutane

To a refluxing solution of triethyl 1,1,2-ethanetricarboxylate (46 ml,200 mmol) and sodium borohydride (20 g, 530 mmol) in t-butanol (400 ml),methanol was added in 3 aliquots over 30 minutes (total 25 ml). Thesolution was refluxed for a further 30 minutes and allowed to cool.Hydrochloric acid (5M) was carefully added to neutralise the solution.The solution was filtered and the inorganic residue was extracted withethanol (2×100 ml) and filtered. The organic solutions were combined andthe solvent removed. The residue was extracted with ethanol (120 ml) andthe solution filtered. The solvent was removed to afford1,4-dihydroxy-2-hydroxy-methylbutane (24 g, 100%); δ_(H) (D₂O) 1.53 (2H,q, J 6 Hz, 3-H), 1.75 (1H, m, 2-H), 3.57 (4H, d, J 6 Hz, 1-H and 1′-H),and 3.64 (2H, t, J 6 Hz, 4-H).

B. 5-(2-Hydroxyethyl)-2,2-dimethyl-1,3-dioxan

To a solution of 1,4-dihydroxy-2-hydroxymethylbutane (12 g, 100 mmol) intetrahydrofuran (25 ml), 2,2-dimethoxypropane (13.5 g, 110 mmol) andp-toluenesulphonic acid monohydrate (0.57 g, 3 mmol) were added. Thesolution was stirred for 0.5 hour at room temperature and was thenneutralised by addition of triethylamine. The solvent was removed andthe residue purified by column chromatography on silica gel eluting withchloroform-methanol mixtures to afford5-(2-hydroxyethyl)-2,2-dimethyl-1,3-dioxan as a clear colourless liquid(6.5 g, 41%).

EXAMPLE 2 5-(2-Bromoethyl)-2,2-dimethyl-1,3-dioxan

To an ice-cooled solution of 5-(2-hydroxyethyl)-2,2-dimethyl-1,3-dioxan(1.92 g, 12 mmol) and carbon tetrabromide (7.96 g, 24 mmol) indimethylformamide (100 ml), triphenyl-phosphine (6.30 g, 24 mmol) wasadded and the solution was left at 4° C. overnight. To this solutionmethanol (20 ml) was added and the solvent was then evaporated underreduced pressure. The residue was purified by column chromatography onsilica gel, eluting with hexane-acetone (12:1) to afford5-(2-bromoethyl)-2,2-dimethyl-1,3-dioxan as a clear colourless liquid(0.89 g, 40%); νmax (film) 2940, 1370, 1270, 1260, 1200, and 1070 cm⁻¹;δ_(H) (CDCl₃) 1.42 (6H, s, C(CH₃)₂), 1.94 (3H, m, CHCH ₂CH₂Br), 3.43(2H, t, J 7 Hz, CH₂Br), and 3.5-4.1 (4H, m, 2×CH₂O); (Found: C, 42.84;H, 6.93%. C₈H₁₅BrO₂ requires: C, 43.07; H, 6.78%. [M−CH₃] found207.0024; C₇H₁₂BrO₂ requires 207.0021).

Alternative Procedure for Preparation of5-(2-Bromoethyl)-2,2-dimethyl-1-3-dioxan (Example 2)

To an ice-cooled solution of 5-(2-hydroxyethyl)-2,2-dimethyl-1,3-dioxan(6.08 g, 38 mmol) and carbon tetrabromide (18.90 g, 57 mmol) inN,N-dimethylformamide (110 ml), triphenyl-phosphine (14.95 g, 57 mmol)was added. The solution was stirred for 0.5 hour at 0° C. The solutionwas then diluted with saturated aqueous sodium bicarbonate (55 ml)followed by water (55 ml), and was extracted with hexane (2×150 ml). Thecombined organic layers were dried (magnesium sulphate) and the solventremoved. The residue was placed under high vacuum for 2 hours to removebromoform. The residue was taken up in a small amount of hexane,filtered and the solvent removed to afford5-(2-bromoethyl)-2,2-dimethyl-1,3-dioxan (7.40 g, 87%) as a colourlessoil which crystallised on cooling, m.p. ca. 18° C.

EXAMPLE 32-Amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]-purine

To a solution of 5-(2-bromoethyl)-2,2-dimethyl-1,3-dioxan (0.75 g, 3.7mmol) in dry dimethylformamide (12 ml) 2-amino-6-chloropurine (0.68 g,4.0 mmol) and then anhydrous potassium carbonate (0.83, 6.0 mmol) wereadded. The solution was stirred at room temperature for 5 hours and leftat 4° C. overnight. The solution was filtered and the solvent removed.The residue was purified by column chromatography on silica gel, elutingwith chloroform-methanol mixtures (80:1 and 60:1) to afford2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine as awhite ctystalline solid (0.74 g, 64%), m.p. 125-126° C.; λmax (H₂O) 223(ε 28,900), 247 (ε 5,700), and 310 (ε 7,700) nm; νmax (KBr) 3450, 3340,1635, 1615, 1565, 1470, 1410, and 1375 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.26 (3H,s, CH₃), 1.32 (3H, s, CH₃), 1.45-1.85 (3H, m, CHCH ₂CH₂N), 3.51 (2H, dd,J 11 Hz and J 7 Hz, 2×H_(ax)), 3.78 (2H, dd, J 11 Hz and J 4 Hz,2×H_(eq)), 4.05 (2H, t, J 7 Hz, CH₂N), 6.89 (2H, s, D₂O exchangeable,2-NH₂), and 8.38 (1H, s, 8-H); (Found: C, 50.37; H, 5.68; N, 22.22%; M⁺311.1136. C₁₃H₁₈ClN₅O₂ requires C, 50.08; H, 5.82; N, 22.46%; M⁺311.1149).

EXAMPLE 4 9-(4-Hydroxy-3-hydroxymethylbut-1-yl)guanine

2-Amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)-ethyl]purine (0.59g, 1.9 mmol) in hydrochloric acid (1.0M, 4 ml) was stirred at 60° C. for24 hours. The solution was diluted with water and neutralised withAmberlite IR 45 (OH⁻ form). The mixture was filtered, the resin washedwith water and the solvent evaporated under reduced pressure. Theresidue was recrystallised from water to afford9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (238 mg, 49%), m.p.275-277° C.; λmax (H₂O) 253 (ε 11,500) and 270 (shoulder, ε 8,630) nm;νmax (KBr) 3420, 3140, 1690, 1645, and 1605 cm⁻¹; δ_(H) [(CD₃)₂SO]1.3-1.5 (3H, m, CHCH ₂CH₂), 3.42 (4H, d, J 5 Hz, 2×CH₂O), 3.99 (2H, t, J7 Hz, CH₂N), 4.41 (2H, br, D₂O exchangeable, 2×OH), 6.44 (2H, s, D₂Oexchangeable, 2-NH₂), 7.71 (1H, s, 8-H), and 10.55 (1H, br, D₂Oexchangeable, 1-H); (M⁺ found 253.1176. C₁₀H₁₅N₅O₃ requires M⁺253.1175).

Alternative Procedure for Preparation of9-(4-Hydroxy-3-hydroxymethylbut-1-yl)guanine (Example 4)

2-Amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)-ethyl]purine (3.74g, 12 mmol) in hydrochloric acid (2.0M, 12 ml) was heated under refluxfor 1.5 hours. The solution was neutralised with aqueous sodiumhydroxide (10%) and then allowed to cool. The solution was filtered andthe solid washed with water to afford9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine as a white crystallinesolid (2.18 g, 72%), m.p. 275-277° C.; (Found: C, 47.31; H, 6.02; N,27.81%; C₁₀H₁₅N₅O₃ requires C, 47.43; H, 5.97; N, 27.65%).

EXAMPLE 5 Ethyl 4-Benzyloxy-2-ethoxycarbonylbutanoate

Sodium metal (72 g, 3.13 mmol) was dissolved in dry ethanol (1.21) withstirring, then diethyl malonate (477 ml, 3.13 mmol) and potassium iodide(249.5 g, 1.5 mmol) were added. An oil containing benzyl-2-bromoethylether (278.5 g, 1.3 mmol) contaminated with ethylene carbonate (198 g)was then added slowly to the stirred mixture. On completion of theaddition, the reaction mixture was heated under reflux for 18 hours,then poured into ice-water (7.51) and extracted with ether (3×1.61). Theether fractions were combined, dried (MgSO₄), filtered and evaporated togive an oil (550 g). This was vacuum distilled to give ethyl4-benzyloxy-2-ethoxycarbonylbutanoate (313 g, 82%) as a clear oil, b.p.165-180°/2 mm. δ_(H) (CDCl₃) 1.22 (6H, t, 2×CH₃), 2.18 (2H, q, CHCH ₂),3.48 (2H, t, CHCH ₂O), 3.55 (1H, t, CH), 4.12 (4H, q, 2×CO₂CH₂), 4.38(2H, s, OCH₂Ph), 7.21 (5H, s, Ar).

EXAMPLE 6 4-Benzyloxy-2-hydroxymethylbutan-1-ol

To a cooled, stirred suspension of lithium aluminium hydride (103 g, 2.7mol) in dry ether (2.51) under nitrogen was added ethyl4-benzyloxy-2-ethoxycarbonyl-butanoate (362 g, 1.23 mol) over a periodof 3 hours. On completion of the addition, the reaction mixture wasallowed to warm to room temperature, and then heated under reflux for 1hour. It was then re-cooled and the excess lithium aluminium hydridedestroyed by dropwise addition of water (100 ml), 2M sodium hydroxide(100 ml) and water (300 ml). The reaction mixture was filtered, and thefilter cake washed well with chloroform. The filtrate was dried (MgSO₄),filtered, and evaporated to give 4-benzyloxy-2-hydroxymethylbutan-1-olas a clear oil (226 g, 87%). δ_(H) (CDCl₃) 1.35-2.05 (3H, m, CHCH ₂CH₂),3.30-3.80 (8H, m, 3×CH₂O, 2×OH), 4.42 (2H, s, OCH ₂Ph), 7.26 (5H, s,Ar).

EXAMPLE 7 2-Acetoxymethyl-4-benzyloxybut-1-yl Acetate

To a cooled solution of 4-benzyloxy-2-hydroxymethyl-butan-1-ol in drypyridine (1.11) was added acetyl chloride (230 ml, 3.24 mol) over 2hours, the temperature being maintained below 8° C. On completion of theaddition, the reaction mixture was stirred at 5° C. for a further 1hour, then poured into water (41) and extracted with ethyl acetate(1×31, 1×21). The organic extracts were combined and washed with 2Mhydrochloric acid (2×1 l), water (1 l) and brine (1 l), dried (MgSO₄),filtered and evaporated to give a pale yellow oil (300 g).

Vacuum distillation afforded 2-acetoxymethyl-4-benzyloxybut-1-yl acetateas a colourless oil (220 g, 70%) b.p. 160-165°/0.05 mm.

The fraction b.p. 122-160°/0.05 mm (42 g) was purified by columnchromatography on silica gel, elution with ether-hexane 2:3 affordingfurther 2-acetoxymethyl-4-benzyloxybut-1-yl acetate (27 g, 8%). δ_(H)(CDCl₃) 1.68 (2H, q, CHCH ₂CH₂), 2.01 (6H, s, 2×OCOCH₃), 2.19 (1H, m,CH), 3.50 (2H, t, CH ₂OCH₂Ph), 4.03 (4H, d, CH ₂OCOCH₃), 4.43 (2H, s,OCH ₂Ph), 7.24 (5H, s, Ar).

EXAMPLE 8 2-Acetoxymethyl-4-hydroxybut-1-yl Acetate

To a solution of 2-acetoxymethyl-4-benzyloxy-but-1-yl acetate (55 g,0.187 mol) in ethanol (250 ml) was added 10% palladium on carbon (2.5g), and the mixture hydrogenated at atmospheric pressure and roomtemperature. When the theoretical hydrogen uptake had been achieved (18hours), the reaction was stopped and filtered through Celite.Evaporation of the filtrate gave a colourless oil (35 g). This waspurified by column chromatography on silica gel, elution with 2%methanol in chloroform affording 2-acetoxy-methyl-4-hydroxybut-1-ylacetate as a clear oil (32.9, 86%). δ_(H) (CDCl₃) 1.61 (2H, q, CHCH₂CH₂), 2.04 (6H, s, 2×OCOCH₃), 2.20 (1H, m, CH), 2.61 (1H, br s, D₂Oexchangeable, OH), 3.68 (2H, t, CH ₂OH), 4.04 (4H, d, 2×CH ₂OCOCH₃).

EXAMPLE 9 2-Acetoxymethyl-4-bromobut-1-yl Acetate

A mixture of 2-acetoxymethyl-4-hydroxy-but-1-yl acetate 10 g, 49 mmol),triphenylphosphine (19.25 g, 73 mmol) and carbon tetrabromide (24.4 g,73 mmol) was stirred for 18 hours at 4° C. in dimethylformamide (150ml). The solvent was then evaporated, and the residue purified by columnchromatography on silica gel, eluting with ether-light petroleum 2:3 toafford 2-acetoxymethyl-4-bromobut-1-yl acetate as a pale oil (130 g,99%). δ_(H) (CDCl₃) 1.73-2.56 (3H, m, CHCH ₂CH₂), 2.04 (6H, s,2×OCOCH₃), 3.44 (2H, t, CH₂Br), 4.04 (4H, d, 2×CH ₂OCOCH₃).

EXAMPLES 11 and 109-(4-Acetoxy-3-acetoxymethylbut-1-yl)-2-amino-6-chloropurine and7-(4-Acetoxy-3-acetoxymethylbut-1-yl)-2-amino-6-chloropurine

A mixture of 2-acetoxymethyl-4-bromobut-1-yl-acetate (13.0 g, 48.7mmol), 2-amino-6-chloro-purine (8.25 g, 48.7 mmol) and anhydrouspotassium carbonate (10 g, 72.5 mmol) was stirred in drydimethylformamide (100 ml) for 18 hours at room temperature. Thereaction mixture was then filtered, the filtrate evaporated, and theresidue purified by column chromatography on silica gel (500 g). Elutionwith 3% methanol in chloroform afforded9-(4-acetoxy-3-acetoxymethyl-but-1-yl)-2-amino-6-chloropurine as a whitesolid (13.8 g, 80%) mp 135-137°. λmax (H₂O) 222 (ε 28,500), 245 (ε4,800) 307 (ε 7,700) nm; νmax (KBr) 3485, 3310, 3200, 1750, 1730, 1625,1560, 1525, 1475, 1245 cm⁻¹. δ_(H) [(CD₃)₂SO] (270 MHz) 1.85-2.05 (3H,m, CHCH ₂CH₂), 2.01 (6H, s, 2×OCOCH₃), 4.03 (4H, d, 2×CH ₂OCOCH₃), 4.16(2H, t, CH₂N), 6.88 (2H, br s, D₂O exchangeable, NH₂), 8.17 (1H, s,8-H). Found C, 47.27; H, 4.94; N, 19.56%. C₁₄H₁₈N₅O₄Cl requires C,47.26; H, 5.10; N 19.68%.

Subsequent elution with 5% methanol in chloroform afforded7-(4-acetoxy-3-acetoxymethylbut-1-yl)-2-amino-6-chloropurine as a whitesolid (2.9 g, 17%) mp 174-175° (dec). λmax (H₂O) 222 (ε 25,000), 255 (ε3,900), 318 (ε 5,600) nm; νmax (KBr) 3390, 3310, 3205, 1745, 1735, 1635,1550, 1505, 1380, 1365, 1310, 1250, 1240 cm⁻¹. δ_(H) [(CD₃)₂SO] (270MHz) 1.86 (2H, q, CHCH ₂CH₂), 1.99 (6H, s, 2×OCOCH₃), 1.95-2.05 (1H, m,CH), 4.03 (4H, d, 2×CH ₂OCCCH₃), 4.38 (2H, t, CH₂N), 6.60 (2H, br s, D₂Oexchangeable, NH₂), 8.39 (1H, s, 8-H). Found C, 47.48; H, 5.11; N,19.52%. C₁₄H₁₈N₅O₄Cl requires C, 47.26; H, 5.10; N, 19.68%.

Alternative Procedure for Preparation of9-(4-Hydroxy-3-hydroxymethylbut-1-yl)guanine (Example 4)

A solution of9-(4-acetoxy-3-acetoxymethyl-but-1-yl)-2-amino-6-chloropurine (15.5 g,43.6 mmol) in 2M hydrochloric acid (150 ml) was heated under reflux for2 hours. The solution was then cooled to room temperature andneutralised with 10% sodium hydroxide solution, left to stand at 4° C.,and the resulting precipitate filtered off, washed with cold water andrecrystallized from water to give9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine as a white crystallinesolid (9.4 g, 85%) mp 275-277°.

EXAMPLE 12 9-(4-Hydroxy-3-hydroxymethylbut-1-yl)guanine, Sodium Salt

To a suspension of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)-guanine (0.30g, 1.2 mmol) in water (8 ml) was added aqueous sodium hydroxide (1M, 1.2ml). The solvent was removed from resulting clear solution andtrituration with methanol-ethanol afforded9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine sodium salt as a whitesolid (0.32 g, 97%); λmax (H₂O, pH7.8) 252 (11,300) nm; νmax (KBr) 3400,1590, and 1570 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.47 (1H, m, 3′-H), 1.70 (2H, q, J7 Hz, 2′-H), 3.3-3.5 (4H, AB part of ABX, 2×4′-H), 3.96 (2H, t, J 7 Hz,1′-H), 4.7 (2H, br, D₂O exchangeable, 2×OH), 5.58 (2H, br.s, D₂Oexchangeable, 2-NH₂), and 7.43 (1H, s, 8-H).

EXAMPLE 13 9-(4-Hydroxy-3-hydroxymethylbut-1-yl)guanine, Potassium Salt

To a suspension of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)-guanine (0.30g, 1.2 mmol) in water (8 ml) was added aqueous potassium hydroxide (1M,1.2 ml). The solvent was removed from the resulting clear solution andtrituration with methanol-ethanol afforded9-(4-hydroxy-3-hydroxymethylbut-1-yl)-guanine potassium salt as a whitepowdery solid (0.34 g, 97%); λmax (H₂O, pH7.4) 252 (12,400); νmax (KBr)3360, 3180, 1690, 1650, 1585, 1570, and 1480 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.47(1H, m, 3′-H), 1.69 (2H, q, J 7.1 Hz, 2′-H), 3.3-3.5 (4H, AB part ofABX, 2×4′-H), 3.94 (2H, t, J 7.3 Hz, 1′-H), 4.7 (2H, br, D₂Oexchangeable, 2×OH), 5.69 (2H, br.s, D₂O exchangeable, 2-NH₂), and 7.38(1H, s, 8-H); (Found: C, 41.11; H, 4.91; N, 23.82%; C₁₀H₁₄N₅O₃Krequires: C, 41.22; J, 4.84; N, 24.04%).

EXAMPLE 14 2-Amino-6-chloro-9-(4-hydroxy-3-hydroxymethylbut-1-yl)purineHydrochloride

To a solution of2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (0.46g, 1.5 mmol) in tetrahydrofuran (4.5 ml), hydrochloric acid (2.0M, 0.5ml) was added. A white precipitate formed and after 0.5 hour thesolution was diluted with further tetrahydrofuran and was filtered togive 2-amino-6-chloro-9-(4-hydroxy-3-hydroxymethylbut-1-yl)purinehydrochloride (290 mg, 63%), decomposed over 165° C.; λmax (H₂O, pH5.5)223 (ε 28,400), 245 (ε 4,620), and 307 (ε 7,620) nm; νmax (KBr) 3370,3330, 3200, 2500, 1650, 1630, 1595, and 1505 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.53(1H, m, CHCH₂CH₂), 1.83 (2H, q, J 7 Hz, CHCH ₂CH₂), 3.35 (4H, d, J 6 Hz,2×CH₂O), 4.19 (2H, t, J 7 Hz, CH₂N), 5.85 (9H, s, D₂O exchangeable,2×OH, NH₂, HCl and H₂O), and 8.57 (1H, s, 8-H). (Found: C, 39.04; H,4.85; N, 22.36%; C₁₀H₁₄ClN₅O₄.HCl requires C, 38.98; H, 4.91; N,22.73.%).

EXAMPLE 15 2-Amino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)-6-methoxypurine

To a solution of2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (0.28g, 0.9 mmol) in methanol (2.5 ml), methanolic sodium methoxide (1M, 1.0ml) was added and the solution was stirred at 50° for 1.5 hours. Thesolution was allowed to cool and hydrochloric acid (5M, 0.2 ml) andwater (0.4 ml) were added. After 15 minutes the solution was neutralisedwith 10% aqueous sodium hydroxide. Silica gel was added and the solventremoved. Column chromatography on silica gel eluting withchloroform-methanol mixtures afforded2-amino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)-6-methoxypurine (185 mg,77%), m.p. 117-119° C.; λmax (H₂O) 213 (ε 22,100), 249 (ε 6,860), and280 (ε 8,410) nm; νmax (KBr) 3400, 3240, 3210, 1640, 1610, 1590, 1410,and 1395 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.47 (1H, m, CHCH₂CH₂), 1.74 (2H, q, J 7Hz, CHCH ₂CH₂), 3.40 (4H, d, J 6 Hz, 2×CH₂O), 3.95 (3H, s, OCH₃), 4.06(2H, t, J 7 Hz, CH₂N), 4.4 (2H, br, D₂O exchangeable, 2×OH), 6.33 (2H,s, D₂O exchangeable, 2-NH₂), and 7.46 (1H, s, 8-H) (Found: M⁺ 267.1340;C₁₁H₁₇N₅O₃ requires M⁺ 267.1331).

EXAMPLE 16 2-Amino-6-ethoxy-9-(4-hydroxy-3-hydroxymethylbut-1-yl)purine

To a suspension of2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (0.31g, 1.0 mmol) in ethanol (1.5 ml) was added sodium ethoxide (1M inethanol, 1.5 ml) and the mixture was stirred at 60° for 1 hour. Theresulting solution was allowed to cool, hydrochloric acid (5M, 0.3 ml)and water (0.7 ml) were added and the solution was stirred for 1 hour atroom temperature. The solution was neutralised by addition of aqueoussodium bicarbonate and the solvent was removed. The residue wasextracted with chloroform-ethanol (2:1), the solution was filtered andthe solvent removed. The residue was purified by column chromatographyon silica gel eluting with chloroform-methanol (6:1) to afford2-amino-6-ethoxy-9-(4-hydroxy-3-hydroxymethylbut-1-yl)purine (0.24 g,85%), m.p. 150-152° C.; λmax (H₂O) 213 (24,300), 249 (7,360), and 280(9,270) nm; νmax (KBr) 3330, 3210, 2900, 1650, 1610, 1580 cm⁻¹; δ_(H)[(CD₃)₂SO] 1.3-1.6 (4H, m, 3′-H and CH₃), 1.73 (2H, q, J 7 Hz, 2′-H),3.2-3.6 (4H, AB part of ABX, 2×4′-H), 4.04 (2H, t, J 7 Hz, 1′-H),4.3-4.55 (4H, m, 2H D₂O exchangeable, 2×OH; D₂O exchange leaves 2H, q, J7 Hz, 6-OCH₂), 6.30 (2H, s, D₂O exchangeable, 2-NH₂), and 7.84 (1H, s,8-H); (Found: C, 50.91; H, 7.00; N, 24.89%; C₁₂H₁₉N₅O₃ requires C,51.23; H, 6.81; N, 24.90%).

EXAMPLE 172-Amino-6-benzyloxy-9-(4-hydroxy-3-hydroxymethylbut-1-yl)purine

A suspension of2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (0.31g, 1.0 mmol) in a solution of sodium benzoxide (1M in benzyl alcohol, 2ml) was stirred at 70° for 1 hour. The resulting solution was allowed tocool, hydrochloric acid (5M, 0.4 ml) and water (0.6 ml) were added andthe solution was stirred for 1 hour at room temperature. The solutionwas then partitioned between chloroform and water. The aqueous layer wasneutralised with aqueous sodium bicarbonate and extracted withchloroform. The combined organic layers were washed with aqueous sodiumbicarbonate, dried (magnesium sulphate) and the solvent was removed. Theresidue was purified by column chromatography on silica gel eluting withchloroform-methanol mixtures (10:1, 5:1) to afford2-amino-6-benzyloxy-9-(4-hydroxy-3-hydroxymethylbut-1-yl)purine as awhite crystalline solid (0.17 g, 50%), m.p. 146-147.5° C.; λmax (EtOH)212 (32,300), 250 (8,380), and 283 (10,100) nm; νmax (fBr) 3340, 3220,1655, 1605, and 1580 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.3-1.6 (1H, m, 3′-H), 1.72(2H, q, J 7 Hz, 2′-H), 3.38 (4H, AB part of ABX, 2×4′-H), 4.03 (2H, t, J7 Hz, 1′-H), 4.36 (2H, t, J 5.5 Hz, D₂O exchangeable, 2×OH), 5.47 (2H,s, PhCH ₂), 6.37 (2H, s, D₂O exchangeable, 2-NH₂), 7.3-7.6 (5H, m,C₆H₅), and 7.84 (1H, s, 8-H); (Found: C, 58.89; H, 6.12; N, 19.87%;C₁₇H₂₁N₅O₃ requires C, 59.46; H, 6.16; N, 20.40%).

EXAMPLE 18 2-Amino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)-6-thiopurine

A solution of2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (0.31g, 1.0 mmol) in aqueous sodium hydrosulphide (2M, 3.0 ml) and ethanol(1.5 ml) was stirred at 70° C. for 1 hour. To this solution glacialacetic acid (2.5 ml) was added and the mixture was stirred for a further1 hour at 70° C. The solution was allowed to cool, filtered and thesolvent removed. The residue was recrystallised from water to afford2-amino-9-(4-hydroxy-3-hydroxy-methylbut-1-yl)-6-thiopurine (0.13 g,48%), m.p. decomposed at 260° C.; δ_(H) [(CD₃)₂SO] 1.45 (1H, m,CHCH₂CH₂), 1.72 (2H, q, J 7 Hz, CHCH ₂CH₂), 3.3-3.5 (4H, ABX J_(AB) 10.7Hz, J_(AX) 5.5 Hz and J_(BX) 5.8 Hz, 2×CH₂O), 4.02 (2H, t, J 7.4 Hz,CH₂N), 4.45 (2H, br, D₂O exchangeable, 2×OH), 6.77 (2H, S, D₂Oexchangeable, 2-NH₂), 7.87 (1H, s, 8-H), and 11.9 (1H, br, D₂Oexchangeable, 1-H); λmax (H₂O) 230 (ε 16,900), 263 (ε 7,210) and 341 (ε25,200) nm; νmax (KBr) 3310, 3130, 1650, 1610, and 1580 cm⁻¹; (Found: C,44.86; H, 5.60; N, 25.44%; C₁₀H₁₅N₅O₂S requires C, 44.60; H, 5.61; N,26.00%).

EXAMPLE 192-Amino-6-azido-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine

To a solution of2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (0.47g, 1.5 mmol) in dry N,N-dimethylformamide (5 ml), sodium azide (0.20 g,3.0 mmol) was added and the mixture was stirred at 100-110° C. for 4hours. The solvent was removed and the residue washed with water toleave 2-amino-6-azido-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine asa crystalline solid (0.36 g, 75%), m.p. decomposed at 200° C.; λmax(MeOH) 272 (ε 8,210) and 301 (ε 10,100) nm; νmax (KBr) 1670, 1625, and1560 cm⁻¹; δ_(H) (CDCl₃—CD₃OD) 1.44 (6H, s, C(CH₃)₂), 1.6-2.2 (3H, m,CHCH ₂CH₂), 3.5-3.8 (2H, dd (ABX), J 7 Hz and J 11 Hz, 2×H_(ax)),3.85-4.15 (2H, dd (ABX), J 4 Hz and J 11 Hz, 2×H_(eq)), 4.29 (2H, t, J 7Hz, CH₂N), and 7.93 (1H, s, 8-H) (Found: C, 48.96; H, 5.66; N, 35.15%;M⁺ 318.1553. C₁₃H₁₈N₈O₂ requires C, 49.05; H, 5.07; N, 35.20%; M⁺318.1546).

EXAMPLE 20 2,6-Diamino-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine

A mixture of2-amino-6-azido-9-[2-(2,2-dimethl-1,3-dioxan-5-yl)ethyl]purine (318 mg,1.0 mmol), formic acid (0.15 ml, 4.0 mmol), concentrated ammonia (0.22ml, 4.0 mmol), 10% palladium-on-charcoal (30 mg) and methanol (10 ml)was heated under reflux for 1 hour. The solution was allowed to cool,filtered and the solvent removed. The residue was purified by columnchromatography on silica gel eluting with chloroform-methanol mixtures(20:1 and 15:1) to give2,6-diamino-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (190 mg,65%), m.p. 202-204° C.); νmax (KBr) 1670, 1640, 1595, and 1410 cm⁻¹;δ_(H) (CDCl₃—CD₃OD) 1.42 (6H, s, C(CH₃)₂), 1.6-2.0 (3H, m, CHCH ₂CH₂),3.5-4.2 (6H, m, 2×CH₂O and CH₂N), and 7.68 (1H, s, 8-H) (Found: C,52.88; H, 6.78; N, 28.36%; M⁺ 292.1652. C₁₃H₂₀N₆O₂ requires C, 53.41; H,6.90; N, 28.75%; M⁺ 292.1648).

EXAMPLE 21 2,6-Diamino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)purine

A solution of2,6-diamino-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (180 mg, 0.6mmol) in 70% acetic acid (10 ml) was stirred for 1 hour at roomtemperature. The solvent was removed, the residue was suspended inmethanol and sodium methoxide was added to neutralise. Columnchromatography on silica gel eluting with chloroform-methanol mixtures(6:1, 4:1 and 3:1) gave2,6-diamino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)purine (95 mg, 63%),m.p. 187-190° C.; λmax (H₂O, pH6.5) 215 (ε 25,500), 255 (ε 7,290), and280 (ε 9,170) nm; νmax 3150, 1680, 1650, 1605, 1590, and 1410 cm⁻¹;δ_(H) [(CD₃)₂SO] 1.46 (1H, m, CHCH₂CH₂) 1.73 (2H, q, J 7.1 Hz, CHCH₂CH₂), 3.3-3.5 (4H, ddd (ABX), J_(AB) 10.6 Hz, J_(AX) 5.5 Hz and J_(BX)5.9 Hz, 2×CH₂O), 4.01 (2H, t, J 7.3 Hz, CH₂N), 4.41 (2H, br, D₂Oexchangeable, 2×OH), 5.70 (2H, s, D₂O exchangeable, NH₂), 6.56 (2H, s,D₂O exchangeable, NH₂), and 7.69 (1H, s, 8-H) (Found: C, 46.09; H, 6.32;N, 31.69%; C₁₀H₁₆N₆O₂.0.1CHCl₃ requires C, 45.91; H, 6.14; N, 31.81%).

EXAMPLE 22 9-(4-Acetoxy-3-acetoxymethylbut-1-yl)guanine

A mixture of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (253 mg, 1.0mmol), 4-dimethylaminopyridine (25 mg) and acetic anhydride (8.5 ml) wasstirred for 4 days at room temperature. The acetic anhydride was removedunder reduced pressure. The residue was purified by columnchromatography on silica gel eluting with chloroform-methanol mixtures(20:1 and 10:1) to afford 9-(4-acetoxy-3-acetoxymethylbut-1-yl)guanine(160 mg, 47%) which was recrystallised from methanol, m.p. 202-205° C.;νmax (KBr) 1737, 1690, 1628, 1600, and 1240 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.79(2H, q, J 6.7 Hz, CHCH ₂CH₂), 1.91 (1H, m, CHCH₂CH₂), 2.00 (6H, s,2×CH₃), 4.00 (6H, m, 2×CH₂O and CH₂N), 6.42 (2H, S, D₂O exchangeable,2-NH₂), 7.71 (1H, s, 8-H), and 10.54 (1H, s, D₂O exchangeable, 1-H);δ_(C) [(CD₃)₂SO] 20.71 (2×CH₃), 28.29 (C-2′), 34.54 (C-3′), 40.59(C-1′), 63.58 (2×C-4′), 116.64 (C-5), 137.58 (C-8), 151.24 (C-4), 153.38(C-2), 156.87 (C-6), and 170.57 (2×COO) (Found: C, 49.62; H, 5.70; N,20.51%; M⁺ 337.1392; C₁₄H₁₉N₅O₅ requires C, 49.85; H, 5.68; N, 20.76%;M⁺ 337.1386).

EXAMPLES 23 and 249-(4-Propionyloxy-3-propionyloxymethylbut-1-yl)guanine (Example 23) andN²-Propionyl-9-(4-propionyloxy-3-propionyloxymethyl-but-1-yl)guanine(Example 24)

A mixture of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (253 mg, 1.0mmol), 4-dimethylaminopyridine (30 mg), propionic anhydride (8 ml) andN,N-dimethylformamide (15 ml) was stirred at room temperature for 66hours. The solvent was removed and the residue subjected to columnchromatography on silica gel eluting with chloroform-methanol mixtures(30:1, 20:1, 10:1). The first compound to elute wasN²-propionyl-9-(4-propionyloxy-3-propionyloxymethylbut-1-yl)guanine (200mg, 47%) which was recrystallised from ether-methanol, m.p. 152-154° C.;νmax (KBr) 1740, 1675, 1610, 1560, and 1185 cm⁻¹; δ_(H) (CDCl₃) 1.14(6H, t, J 7.5 Hz, 2×OCOCH₂CH ₃), 1.27 (3H, t, J 7.5 Hz, NCOCH₂CH ₃),1.88 (2H, q, J 6.9 Hz, CHCH ₂CH₂), 2.01 (1H, m, CHCH₂CH₂), 2.35 (2H, q,J 7.5 Hz, 2×OCOCH ₂CH₃), 2.56 (2H, q, J 7.5 Hz, NCOCH ₂CH₃), 4.1-4.3(6H, m, 2×CH₂O and CH₂N), 7.65 (1H, s, 8-H), 9.14 (1H, s, N—H) and 11.95(1H, s, N—H) (Found: C, 54.13; H, 6.44; N, 16.19%; M⁺ 421.1958;C₁₉H₂₇N₅O₆ requires C, 54.15; H, 6.46; N, 16.62%; M⁺ 421.1961).

The second compound to elute was9-(4-propionyloxy-3-propionyloxymethylbut-1-yl)guanine (150 mg, 41%)which was recrystallised from methanol, m.p. 204-206° C.; νmax (KBr)3310, 3150, 1740, 1690, and 1190 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.01 (6H, t, J7.4 Hz, 2×CH₂CH ₃), 1.80 (2H, q, J 7.0 Hz, CHCH ₂CH₂), 1.91 (1H, m,CHCH₂CH₂), 2.29 (4H, q, J 7.5 Hz, 2×CH ₂CH₃), 4.01 (6H, m, 2×CH₂O andCH₂N), 6.37 (2H, s, D₂O exchangeable, 2-NH₂), 7.69 (1H, s, 8-H), and10.50 (1H, s, D₂O exchangeable, 1-H) (Found: C, 52.28; H, 6.20; N,18.95%; C₁₆H₂₃N₅O₅ requires C, 52.59; H, 6.35; N 19.17%).

EXAMPLE 25 9-(4-Hexanoyloxy-3-hexanoyloxymethylbut-1-yl)guanine

A mixture of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (253 mg, 1.0mmol), dicyclohexylcarbodiimide (0.83 mg, 4.0 mmol), hexanoic acid (0.38ml, 0.35 g, 3.0 mmol), 4-dimethylaminorpridine (20 mg) andN,N-dimethylformamide (5 ml) was stirred for 64 hours at roomtemperature. The mixture was diluted with water and extracted withchloroform (×2). The combined organic layers were washed with aqueoussodium bicarbonate, dried (magnesium sulphate) and the solvent removed.The residue was purified by column chromatography eluting withchloroform-methanol mixtures to afford9-(4-hexanoyloxy-3-hexanoyloxymethylbut-1-yl)guanine (200 mg, 45%) whichwas recrystallised from menthanol, m.p. 198.5-201° C.; νmax (KBr) 3340,3160, 2960, 2930, 1740, 1690, 1650, 1605, and 1170 cm⁻¹; δ_(H) (CDCl₃)0.87 (6H, t, J 6.9 Hz, 2×CH₃), 1.28 (8H, m, 2×CH ₂CH ₂CH₃), 1.60 (4H,quintet, J 7.4 Hz, 2×COCH₂CH ₂), 1.90 (2H, q, J 6.9 Hz, CHCH ₂CH₂N),2.02 (1H, m, CHCH₂CH₂N), 2.30 (4H, t, J 7.6 Hz, 2×COCH ₂CH₂), 4.13 (6H,m, 2×CH₂O and CH₂N), 6.42 (2H, s, D₂O exchangeable, 2-NH₂), 7.70 (1H, s,8-H), and 12.16 (1H, s, D₂O exchangeable, 1-H) (Found: C, 53.97; H,7.92; N15.45%; C₂₂H₃₅N₅O₅ requires C, 58.78; H, 7.85; N, 15.58%).

EXAMPLE 26 9-(4-Formyloxy-3-formyloxymethylbut-1-yl)guanine

A mixture of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (0.23 g, 0.9mmol), dicyclohexylcarbodiimide (0.92 g, 4.5 mmol), formic acid (0.17ml, 4.5 mmol), 4-dimethylaminopyridine (20 mg) and N,N-dimethylformamide(5 ml) was stirred for 40 minutes at room temperature and then quenchedby addition of methanol (1 ml). The solution was filtered and thesolvent removed. The residue was purified by column chromatography onsilica gel eluting with chloroform-methanol mixtures (7:1, 4:1) toafford 9-(4-formyloxy-3-formyloxymethylbut-1-yl)guanine which wascrystallised from methanol (0.12 g, 43%), m.p. 195-198° C.; νmax (KBr)1720, 1680, 1630, 1600, and 1570 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.83 (2H, q, J7.1 Hz, 2′-H), 2.01 (1H, m 3′-H), 4.04 (2H, t, J 7.1 Hz, 1′-H), 4.13(4H, d, J 5.5 Hz, 2×4′-H), 6.39 (2H, s, D₂O exchangeable, 2-NH₂), 7.70(1H, s, 8-H), 8.23 (2H, s, 2×HCOO), and 10.52 (1H, s, D₂O exchangeable,1-H); (Found: C, 45.40; H, 4.68; N, 21.70%; C₁₂H₁₅N₅O₅ requires: C,46.60; H, 4.89; N, 22.64%).

EXAMPLE 279-[4-(N-Imidazolylcarbonyloxy)-3-(N-imidazolylcarbonyloxymethyl)-but-1-yl]guanine

A mixture of 9-(4-hydroxy-3-hydroymethylbut-1-yl)guanine (253 mg, 1.0mmol), N,N′-carbonyldiimidazole (187 mg, 1.15 mmol),4-dimethylaminopyridine (20 mg) and N,N-dimethylformamide (5 ml) wasstirred at room temperature. After 2 hours a further quantity ofN,N′-carbonyldiimidazole (180 mg) was added and stirring was continuedfor a further 2 hours. The solvent was removed and the residue washedwith water, ethyl acetate and hot methanol leaving9-[4-(N-imidazolylcarbonyloxy)-3-(N-imidazomylcarbonyloxymethyl)but-1-yl]guanine(360 mg, 82%), m.p. >300° C.; νmax (KBr) 3320, 3140, 1760, 1695, 1630,and 1600 cm⁻¹; δ_(H) [(CD₃)₂SO] 2.0 (2H, q, J 7 Hz, CHCH ₂CH₂),2.15-2.40 (1H, m, CHCH₂CH₂), 4.10 (2H, t, J 7 Hz, CH₂N), 4.48 (4H, d, J5 Hz, 2×CH₂O), 6.32 (1H, s, D₂O exchangeable, 2-NH₂), 7.05 (2H, s,imid-H), 7.57 (2H, s, imid-H), 7.75 (1H, s, 8-H), 8.28 (2H, s, imid-H),and 10.53 (1H, s, D₂O exchangeable, 1-H); M/Z 68 (100, imidazole⁺), 44(70, CO₂ ⁺), 41 (72, NCHN⁺).

EXAMPLE 28 & 29N²-Monomethoxytrityl-9-(4-monomethoxytrityloxy-3-hydroxy-methylbut-1-yl)guanineand N²-Monomethoxytrityl-9-(4-hydroxy-3-hydroxymethylbut-1-yl)-guanine

A solution of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)-guanine (4.05 g, 16mmol), monomethoxytrityl chloride (10.9 g 35 mmol), triethylamine (6.7ml) and 4-dimethylaminopyridine (40 mg) in N,N-dimethylformamide (50 ml)was stirred for 2 hours. The reaction was quenched with methanol and thesolvent was removed. The residue was taken up in ethyl acetate and thesolution washed with aqueous sodium bicarbonate and water. The solutionwas dried (magnesium sulphate) and the solvent removed. The residue waspurified by column chromatography on silica gel eluting withchloroform-methanol mixtures. The first major product to elute wasN²-monomethoxytrityl-9-(4-monomethoxytrityloxy-3-hydroxymethylbut-1-yl)guanine(4.4 g, 34%), m.p. 142-145° C.; λmax (EtOH) 230 (sh. 29,900) and 262(16,000) nm; νmax (KBr) 3400, 1680, 1605, 1570, and 1510 cm⁻¹; δ_(H)[(CD₃)₂SO] 1.24 (2H, m, 2′-H), 1.43 (1H, m, 3′-H), 2.7-2.9 (2H, AB partof ABX, CH₂OC), 3.1-3.4 (2H, AB part of ABX CH ₂OH), 3.42 (2H, t, J 6.7Hz, 1′-H), 3.66 (3H, s, CH₃O), 3.74 (3H, s, CH₃O), 4.35 (1H, t, J 4.8Hz, D₂O exchangeable, OH), 6.7-7.4 (28H, m, Ar—H), 7.44 (1H, s, 8-H),7.55 (1H, s, D₂O exchangeable, 2-NH), and 10.50 (1H, s, D₂Oexchangeable, 1-H); (Found: C, 74.28; H, 5.86; N, 8.64%; C₅₀H₄₇N₅O₅requires: C, 75.26; H, 5.94; N, 8.78%).

The second major product to elute wasN²-monomethoxytrityl-9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (1.4g, 17%), m.p. 205-207° C.; λmax (EtOH) 261 (14,500) nm; νmax (KBr) 3380,1705, 1680, 1610, 1570, and 1515 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.25 (3H, m,2′-H and 3′-H), 3.1-3.3 (4H, m, 2×4′-H), 3.52 (2H, t, J 6.6 Hz, 1′-H),3.72 (3H, s, CH₃O), 4.28 (2H, t, J 5.2 Hz, D₂O exchangeable, 2×OH),6.85-7.35 (14H, m, Ar—H), 7.54 (1H, s, 8-H), 7.56 (1H, s, D₂Oexchangeable, 2-NH), and 10.49 (1H, s, D₂O exchangeable, 1-H); (Found:C, 67.93; H, 6.05; N, 12.90% C₃₀H₃₁N₅O₄ requires C, 68.55; H, 5.95; N,13.32%).

EXAMPLE 30 9-(4-Pivalyloxy-3-pivalyloxymethylbut-1-yl)guanine

To a solution ofN²-monomethoxytrityl-9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (0.47g, 0.9 mmol) in pyridine (4.5 ml) was added pivalyyl chloride (0.55 ml,4.5 mmol) and the solution was stirred for 45 minutes. The mixture wasprecipitated in water (45 ml) and the resulting precipitate was stirredin 80% acetic acid (10 ml) at 80° for 20 minutes. The solvent wasremoved and the residue was purified by column chromatography on silicagel eluting with chloroform-methanol (10:1) to afford9-(4-pivalyloxy-3-pivalyloxy-methylbut-1-yl)guanine (0.22 g, 58%), m.p.225-237° C.; νmax (KBr) 3430, 2980, 1730, 1690, and 1620 cm⁻¹; δ_(H)[(CD₃)₂SO] 1.11 (18H, s, 2×C(CH₃)₃), 1.81 (2H, q, J 6.8 Hz, 2′-H), 1.93(1H, m, 3′-H), 4.0-4.1 (6H, m, 1′-H and 2×4′-H), 6.38 (2H, s, 2-NH₂),7.69 (1H, s, 8-H), and 10.58 (1H, br.s, 1-H); (Found: C, 56.58; H, 7.26;N, 16.14%; C₂₀H₃₁N₅O₅ requires: C, 56.99; H, 7.41; N, 16.62%).

EXAMPLE 31 9-(4-Acetoxy-3-hydroxymethylbut-1-yl)guanine

To a solution ofN²-monomethoxytrityl-9-(4-monomethoxytrityloxy-3-hydroxymethylbut-1-yl)guanine(0.72 g, 0.9 mmol) in pyridine (3 ml) was added acetyl chloride (0.21ml, 3.0 mmol) and the solution was stirred for 30 minutes. The mixturewas precipitated in water (30 ml) the resulting precipitate was stirredin 80% acetic acid (10 ml) at 80° for 30 minutes. The solvent wasremoved and the residue purified by column chromatography on silica geleluting with chloroform-methanol mixtures (7:1, 3:1) to afford9-(4-acetoxy-3-hydroxymethylbut-1-yl)guanine (0.17 g, 64%), m.p.194-200° C.; νmax (KBr) 3330, 3170, 2930, 1730, 1690, 1660, 1610, and1565 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.6-1.8 (3H, m, 2′-H and 3′-H), 1.98 (3H, s,CH₃), 3.39 (2H, br, D₂O exchange gives d, J 5 Hz, CH ₂OH), 3.9-4.1 (4H,m, 1′-H and CH₂OCO), 4.61 (1H, br.t, D₂O exchangeable, OH), 6.44 (2H, s,D₂O exchangeable, 2-NH₂), 7.68 (1H, s, 8-H), and 10.59 (1H, s, D₂Oexchangeable, 1-H); (Found: C, 47.91; H, 5.63; N, 21.71%; C₁₂H₁₇N₅O₄requires: C, 48.81; H, 5.80; N, 23.72%).

EXAMPLE 32 9-(4-Benzoyloxy-3-hydroxymethylbut-1-yl)guanine

To a solution ofN²-monomethoxytrityl-9-(4-monomethoxy-trityloxy-3-hydroxymethylbut-1-yl)guanine(0.72 g, 0.9 mmol) in pyridine (4 ml) was added benzoyl chloride (0.31ml, 2.7 mmol) and the solution was stirred for 30 minutes. The mixturewas precipitated in water (40 ml) and the resulting precipitate wasstirred in 80% acetic acid (10 ml) at 80° for 45 minutes. The solventwas removed and the residue purified by column chromatography on silicagel eluting with chloroform-methanol mixtures (6:1, 3:1) to afford9-(4-benzoyloxy-3-hydroxymethylbut-1-yl)guanine (80 mg, 25%), m.p.156-168° C.; λmax (MeOH) 231 (15,100) and 254 (13,100) nm; νmax (KBr)1715, 1690, 1625, and 1600 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.75-1.90 (3H, m, 2′-Hand 3′-H), 3.50 (2H, t, J 5 Hz, D₂O exchange leaves d, CH ₂OH), 4.07(2H, t, J 6.9 Hz, 1′-H), 4.2-4.35 (2H, AB part of ABX, CH₂OCO), 4.68(1H, t, J 5.1 Hz, D₂O exchangeable, OH), 6.39 (2H, s, D₂O exchangeable,2-NH₂), 7.5-8.0 (6H, m, C₆H₅ and 8-H), and 10.53 (1H, s, D₂Oexchangeable, 1-H); (Found: C, 53.57; H, 5.28; N, 17.95%;C₁₇H₁₉N₅O₄.0.25 CHCl₃ requires: C, 53.51; H, 5.01; N, 18.09%).

EXAMPLE 33 9-(4-Hexanoyloxy-3-hydroxymethylbut-1-yl)guanine

To a solution ofN²-monomethoxytrityl-9-(4-monomethoxy-trityloxy3-hydroxymethylbut-1-yl)guanine(0.72 g, 0.9 mmol) in pyridine (4 ml) was added hexanoyl chloride (0.38ml, 2.7 mmol) and the solution was stirred for 20 minutes. The mixturewas precipitated in water (40 ml) and the resulting precipitate wasstirred in 80% acetic acid (10 ml) at 80° for 45 minutes. The solventwas removed and the residue purified by column chromatography on silicagel eluting with chloroform-methanol mixtures (7:1, 5:1) to afford9-(4-hexanoyloxy-3-hydroxymethylbut-1-yl)guanine (0.12 g, 38%), m.p.179-181° C.; νmax 2960, 2930, 1730, 1690, 1630, and 1600 cm⁻¹; δ_(H)[(CD₃)₂SO] 0.84 (3H, t, J 6.9 Hz, CH₃), 1.24 (4H, m, CH₃(CH ₂)₂), 1.50(2H, quintet, J 7.3 Hz, CH ₂CH₂CO), 1.6-1.8 (3H, m, 2′-H and 3′-H), 2.26(2H, t, J 7.3 Hz, CH₂CO), 3.40 (2H, t, J 5 Hz, D₂O exchange leaves d, CH₂OH), 3.9-4.1 (4H, m, 1′-H and CH₂OCO), 4.60 (1H, t, J 5.1 Hz, D₂Oexchangeable, 2-NH₂), 7.67 (1H, s, 8-H), and 10.49 (1H, s, D₂Oexchangeable, 1-H); (Found: C, 52.63; H, 6.91; N, 18.75:%; C₁₆H₂₅N₅O₄requires: C, 54.69; H, 7.17; N, 19.93%).

EXAMPLE 34 9-(4-Hexadecanoyloxy-3-hydroxymethylbut-1-yl)guanine

To a solution ofN²-monomethoxytrityl-9-(4-monomethoxy-trityloxy-3-hydroxymethylbut-1-yl)guanine(0.72 g, 0.9 mmol) in pyridine (4 ml) was added hexadecanoyl chloride(0.82 ml, 2.7 mmol) and the solution was stirred for 30 minutes. Themixture was precipitated in water (40 ml) and the resulting precipitatewas stirred in 80% acetic acid (8 ml) at 80° for 2 hours. The solventwas removed and the residue was purified by column chromatography onsilica gel eluting with chloroform-methanol mixtures (10:1, 8:1) toafford 9-(4-hexadecanoyloxy-3-hydroxymethylbut-1-yl)guanine (0.23 g,52%), m.p. 183-191° C.; νmax (KBr) 3340, 3160, 2920, 2850, 1740, 1690,and 16.05 cm⁻¹; δ_(H) [(CD₃)₂SO] 0.85 (3H, t, J 6.6 Hz, CH₃), 1.23 (24H,m, CH₃(CH ₂)₁₂), 1.49 (2H, m, CH ₂CH₂CO), 1.6-1.8 (3H, m, 2′-H and3′-H), 2.26 (2H, t, J 7.3 Hz, CH₂CO), 3.39 (2H, t, J 5 Hz, D₂O exchangeleaves d, CH ₂OH), 3.9-4.1 (4H, m, 1′-H and CH₂OCO), 4.60 (1H, t, J 5.2Hz, D₂O exchangeable, OH), 6.38 (2H, s, D₂O exchangeable, 2-NH₂), 7.67(1H, s, 8-H), and 10.50 (1H, s, D₂O exchangeable, 1-H); (Found: C,63.8.3; H, 9.44; N, 14.05%; C₂₆H₄₅N₅O₄ requires: C, 63.51; H, 9.23; N,14.24%).

EXAMPLE 35 9-(4-Hydroxy-3-hydroxymethylbut-1-yl)guanine 4′-PhosphateDiammonium Salt

To a solution of cyanoethyl phosphoric acid (4.8 mmol) in pyridine (6ml) were addedN²-monomethoxytrityl-9-(4-monomethoxytrityloxy-3-hydroxymethylbut-1-yl)guanine(1.28 g, 1.6 mmol) and dicyclohexylcarbodiimide (1.98 g, 9.6 mmol) andthe solution was stirred for 2 hours. Reaction was quenched by additionof water (1 ml) and the solvent was removed. To the residue was addedconcentrated aqueous ammonia and the mixture was stirred at 60° for 3hours. The solvent was removed and to the residue was added 80% aceticacid (15 ml). The mixture was stirred at 80° for 45 minutes and thesolvent was removed. The residue was taken up in water (25 ml) andextracted with chloroform (4×30 ml). The aqueous layer was filtered,concentrated and passed down a column of XAD-4 resin, eluting withaqueous methanol mixtures. Fractions containing product were pooled andthe solvent removed. The residue was taken up in a small volume of waterand the solution was passed through C₁₈-Sep-pak cartridges. Fractionscontaining product were pooled and the solvent removed to afford9-(4-hydroxy-3-hydroxymethyl-but-1-yl)guanine 4′-phosphate, diammoniumsalt as a white solid (0.31 g, 53%); νmax (KBr) 3150 (broad), 1690, and1610 cm⁻¹; δ_(H) [(CD₃)₂SO/D₂O] 1.57 (1H, m, 3′-H), 1.70 (2H, m, 2′-H),3.37 (2H, d, 4.7 Hz, CH ₂OH), 3.72 (2H, m, CH₂OP), 3.99 (2H, t, J 6.9Hz, 1′-H), and 7.68 (1H, s, 8-H).

EXAMPLE 36 N²-Acetyl-9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine

To a suspension of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (0.23 g,0.9 mmol) in pyridine (3 ml) was added chlorotrimethylsilane (0.25 ml,2.0 mmol) and the mixture was stirred for 15 minutes. To this mixturewas added acetyl chloride (0.085 ml, 1.2 mmol) and the mixture wasstirred for 30 minutes. Methanol (2 ml) was the added and the mixturewas stirred for a further 30 minutes. The solvent was removed and theresidue purified by column chromatography on silica gel eluting withchloroform-methanol mixtures (4:1, 5:2) to give the title compound asits hydrochloride salt. This was dissolved in methanol and stirred withpotassium carbonate. The solution was filtered and the solvent removed.The residue was taken up in water and passed through C₁₈-Sep-pakcartridges. Fractions containing product were pooled and the solventremoved to afford N²-acetyl-9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine(0.11 g, 41%), m.p. 143-146° C.; λmax (H₂O) 260 (15,100) nm; νmax (KBr)3420, 3200, 2940, 1685, 1615, and 1560 cm⁻¹; δ_(H) [(CD₃)₂SO] 1.46 (1H,m, 3′-H), 1.77 (2H, q, J 7.1 Hz, 2′-H), 2.18 (3H, s, CH₃), 3.3-3.5 (4H,m, 2×4′-H), 4.13 (2H, t, J 7.4 Hz, 1′-H), 4.42 (2H, br.t, J 5 Hz, D₂Oexchangeable, 2×OH), 7.98 (1H, s, 8-H), and 11.83 (2H, br, D₂Oexchangeable, 1-H and 2-NH).

EXAMPLE 37 N²-Hexanoyl-9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine

To a suspension of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)-guanine (0.25g, 1.0 mmol) in pyridine (5 ml) was added chlorotrimethylsilane (0.32ml, 2.5 mmol) and the mixture was stirred for 15 minutes. To thismixture was added hexanoyl chloride (0.18 ml, 1.3 mmol) and the mixturewas stirred for 20 minutes. Methanol (2 ml) was then added and themixture was stirred for a further 20 minutes.1,8-Diazabicyclo[5.4.0]undec-7-ene (0.57 ml, 3.8 mmol) and water (0.5ml) were added and the solvent was removed. The residue was purified bycolumn chromatography on silica gel eluting with chloroform-methanol(4:1). Product containing fractions were pooled and the solvent removed.The residue was taken up in water and passed through C₁₈-Sep-pakcartridges to affordN²-hexanoyl-9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine (50 mg, 14%),m.p. 86-88° C.; νmax (KBr) 3400, 2960, 2940, 1675, 1610, and 1560 cm⁻¹;δ_(H) [(CD₃)₂SO] 0.88 (3H, t, J 6.7 Hz, CH₃) 1.29 (4H, m, CH₃(CH ₂)₂),1.46 (1H, m, 3′-H), 1.60 (2H, m, CH ₂CH₂CO), 1.77 (2H, q, J 7.1 Hz,2′-H), 2.46 (2H, t, J 7.4 Hz, CH₂CO) 3.3-3.5 (4H, m, 2×4′-H), 4.13 (2H,t, J 7.4 Hz, 1′-H), 4.41 (2H, t, J 4.7 Hz, D₂O exchangeable, 2×OH), 7.98(1H, s, 8-H), 11.65 (1H, br, D₂O exchangeable, NH), and 12.01 (1H, br,D₂O exchangeable, NH); (Found: C, 53.64; H, 7.56; N, 18.95%;C₁₆H₂₅N₅O₄.0.5H₂O requires: C, 53.32; H, 7.27; N, 19.43%).

EXAMPLE 382-Amino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)-6-isopropoxypurine

A solution of2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (0.25g, 0.8 mmol) in isopropanol (2.5 ml) containing sodium isopropoxide(0.5M) was stirred at 60° for 25 minutes. After cooling, hydrochloricacid (5M, 0.3 ml) and water (0.7 ml) were added and the solution wasstirred for 15 minutes at room temperature. The solution was neutralisedby addition of aqueous sodium bicarbonate and the solvent was removed.The residue was extracted with chloroform-ethanol (2:1) and the solutionpurified by column chromatography on silica gel eluting withchloroform-methanol (7:1) to afford2-amino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)-6-isopropoxypurine whichwas crystallised from chloroform-carbon tetrachloride (0.18 g, 76%),m.p. 111.5-113.5° C.; δ_(H) [(CD₃)₂SO] 1.34 (6H, d, J 6.3 Hz, C(CH₃)₂),1.45 (1H, m, 3′-H), 1.74 (2H, q, J 7.2 Hz, 2′-H), 3.3-3.5 (4H, AB partof ABX, 2×4′-H), 4.06 (2H, t, J 7.3 Hz, 1′-H), 4.40 (2H, br, 2×OH), 5.49(1H, septet, J 6.3 Hz, CH(CH₃)₂), 6.27 (2H, s, 2-NH₂), and 7.83 (1H, s,8-H).

EXAMPLE 39 2-Amino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)-6-phenoxypurine

To a solution of phenol (113 mg, 1.2 mmol) in dry dioxan (2.5 ml) wasadded sodium hydride (60% dispersion in oil; 48 mg, 1.2 mmol). Afterevolution of hydrogen ceased,2-amino-6-chloro-9-[2-(2,2-dimethyl-1,3-dioxan-5-yl)ethyl]purine (0.25g, 0.8 mmol) was added and the mixture was stirred at 75° for 3.5 hours.After cooling, water (0.8 ml) and hydrochloric acid (5M, 0.2 ml) wereadded and the solution was stirred for 30 minutes at room temperature.The solution was neutralised by addition of aqueous sodium bicarbonateand the solvent was removed. The residue was extracted withchloroform-ethanol (2:1) and the solution purified by columnchromatography on silica gel eluting with chloroform-methanol (9:1) toafford 2-amino-9-(4-hydroxy-3-hydroxymethylbut-1-yl)-6-phenoxypurine(145 mg, 55%), m.p. 173-175° C.; δ_(H) [(CD₃)₂SO] 1.48 (1H, m, 3′-H),1.78 (2H, q, J 7.1 Hz, 2′-H), 3.3-3.5 (4H, m, 2×4′-H), 4.12 (2H, t, J7.4 Hz, 1′-H), 4.42 (2H, t, J 5.1 Hz, D₂O exchangeable, 2×OH), 6.35 (2H,s, D₂O exchangeable, 2-NH₂), 7.2-7.5 (5H, m, C₆H₅), and 7.98 (1H, s,8-H).

Example of Pharmaceutical Activity

Method 1

Vero (Arican Green Monkey Kidney) cells were grown to confluence in 24well multidishes, each well being 1.6 cm in diameter. The cells wereinfected with Herpes simplex type 1 virus (HFEM strain) and overlaidwith 0.5 ml of 0.9% agarose (w/v) in maintenance medium. The testcompound, prepared in maintenance medium in concentrations ranging from100 to 0.3 μg/ml in half-log dilution steps, was added in 0.5 ml volume.The virus infected cultures were then incubated at 37° C. for 4 daysbefore fixing in 4% formaldehyde solution and staining with carbolfuchsin. The dishes were then examined to find what concentration oftest compound caused a 50% reduction in the number of virus plaquesformed (PDD₅₀ value) and the minimum concentration of test compoundwhich caused cytotoxicity (MTD).

Method 2

MRC-5 cells were infected in suspension with Herpes simplex type 1virus, strain SC16. The infected cell suspension was dispensed (0.1 ml)in 96 well microtitre plates containing the test drugs in maintenancemedium in concentrations ranging from 100 to 0.03 μg/ml in half-logdilution steps (0.1 ml per well). The plates were then icubated at 37°C. for 3 days when the virus cytopathic effect (CPE) in the controlwells reached 100%. The plates were fixed in 4% formaldehyde solutionand stained with carbol fuchsin. The plates were then examined to findwhat concentration of test compound reduced the virus CPE by 50% (IC₅₀).Plates using uninfected cells were run in parallel to determine theminimum concentration of test compound which caused cytotoxicity (MTD).

Compounds were also tested against Herpes simplex type 2 virus (MSstrain) in Vero cells using Method 1 and in MRC-5 cells using Method 2.In the latter test, the incubation time was reduced to 24 hours.

Results PDD₅₀ (μg/ml) Herpes simplex type 1 virus Herpes simplex type 2virus HFEM strain SC16 strain MS strain MS strain in Vero in MRC-5 inVero in MRC-5 Example No. cells cells cells cells 4 1.3 0.9 2.3 0.6 122.2 0.7 13 1.7 0.7 15 >100 >100 63 49 22 >100 >100 >100 83 23 >10025 >100 85 24 >100 >100 >100 65 25 1.9 1.0 1.6 0.9 27 13 1.5 2.8 5.7 3124 10 32 95 3 34 16 2

None of the compounds was cytotoxic at concentrations up to 100 μg/ml inany of the tests.

Method 3

Compounds were administered by oral gavage (0.2 mmoles/kg in 0.1 ml of1% carboxymethyl cellulose) to 20 g female Balb/C mice which had beenstarved for 18 hours. Fifteen minutes later, blood was collected fromthree mice by cardiac puncture using heparinised syringes. Equalaliquots were pooled and an equal volume of 16% trichloroacetic acidadded. Following centrifugation (8,500 g) to remove precipitatedproteins, the resulting mixture was analysed by high performance liquidchromatography using a C₁₈ Nova-Pak cartridge eluted with Buffer A (50mM NaH₂PO₄, pH4.6) and Buffer B (10% Buffer A, 10% water, 80% methanol)in a gradient from 1% to 95% Buffer B.9-(4-Hydroxy-3-hydroxymethylbut-1-yl)guanine was assayed with aPye-Unicam PU4021 u.v. detector set at 254 nm.

Results

Concentrations of 9-(4-Hydroxy-3-hydroxymethylbut-1-yl)guanine (Example4) in the Blood of Mice After Oral Administration of Derivatives

Concentration of Example 4 Administered compound in Blood (μg/ml)Example 15 0.2 Example 16 2.3 Example 17 4.8 Example 22 2.0 Example 232.5 Example 30 0.3 Example 35 1.1

What is claimed is:
 1. 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guaninehaving a purity of greater than 95% by weight. 2.9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine having an ¹H-NMR spectrumof d_(H) [(CD₃)₂SO] 1.3-1.5 (3H, m, CHCH₂CH₂), 3.42 (4H, d, J 5 Hz,2×CH₂O), 3.99 (2H, t, J 7 Hz, CH₂N), 4.41 (2H, br, D₂O exchangeable,2×OH), 6.44 (2H, s, D₂O exchangeable, 2-NH₂), 7.71 (1H, s, 8-H), and10.55 (1H, br, D₂O exchangeable, 1-H), and having substantially nodetectable signal in the δ 7.1-7.4 region of the ¹H-NMR spectrum. 3.9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine in substantially pure form.4. The sodium salt of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine, saidsalt being in substantially pure form. 5.9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine having a purity of greaterthan 90% by weight.
 6. The compound9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine, sodium salt, having apurity of greater than 90% by weight.
 7. The compound according to claim6 which has a purity of greater than 95% by weight.
 8. Apharmaceutically acceptable salt of9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine, having a purity of atleast 90% by weight.
 9. A pharmaceutically acceptable salt of9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine, having a purity of atleast 95% by weight.
 10. 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine,in crystalline form, having a purity of greater than 90% by weight. 11.9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine, in crystalline form,having a purity of greater than 95% by weight.
 12. A pharmaceuticallyacceptable salt of 9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine.